Influenza virus protein PB1-F2 interacts with CALCOCO2 (NDP52) to modulate innate immune response

Olivier Leymarie (1) , Léa Meyer (1) , Lionel Tafforeau (2) , Vincent Lotteau (3) , Bruno da Costa (1) , Bernard Delmas (1) , Christophe Chevalier (1) , Ronan Le Goffic (1)

1 VIM (UR 0892) - Unité de recherche Virologie et Immunologie Moléculaires
2 IMAP - Institut des Matériaux Poreux de Paris
3 CIRI - Centre International de Recherche en Infectiologie

PB1-F2 is a viral protein encoded by influenza A viruses (IAVs). PB1-F2 is implicated in virulence by triggering immune cell apoptosis and enhancing inflammation. To obtain an insight into the molecular mechanisms of PB1-F2-mediated virulence, we used the yeast two-hybrid approach to find new PB1-F2 cellular interactors. This allowed us to identify calcium-binding and coiled-coil domain 2 (CALCOCO2, also known as NDP52) as a binding partner of PB1-F2. Binding of PB1-F2 to CALCOCO2 was confirmed by pull-down. Surface plasmon resonance binding experiments enabled us to estimate the dissociation constant (Kd) of the two partners to be around 20 nM. Using bioinformatics tools, we designed a CALCOCO2 interaction map based on previous knowledge and showed a strong connection between this protein and the type I interferon production pathways and the I-\kappaB kinase/NF-\kappaB signalling pathway. NF-\kappaB reporter assays in which CALCOCO2, MAVS and PB1-F2 were co-expressed showed a cooperation of these three proteins to increase the inflammatory response. By contrast, PB1-F2 inhibits the TBK1-dependent activation of an ISRE reporter plasmid. We also demonstrated that the signal transducer TRAF6 is implicated in the enhancement of NF-\kappaB activity mediated by PB1-F2/CALCOCO2 binding. Altogether, this report provides evidence of an interaction link between PB1-F2 and human proteins, and allows a better understanding of the involvement of PB1-F2 in the pathologic process mediated by IAV.

Mots clés: Protein Binding; Protein Interaction Mapping; Surface Plasmon Resonance; Computational Biology; Host-Pathogen Interactions; Humans; Immunity; Innate; Influenza A virus; Kinetics; Nuclear Proteins; Two-Hybrid System Techniques; Viral Proteins; Virulence Factors; PB1-F2; CALCOCO2; calcium-binding and coiled-coil domain 2; IAV; influenza A virus; ISRE; IFN-stimulated response element; MAVS; mitochondrial antiviral signalling protein; qRT-PCR; quantitative reverse transcription PCR; SPR; surface plasmon resonance; siRNA; small interfering RNA; TRAF6; TNF receptor-associated factor 6; TRAF6 DN; dominant-negative form of TRAF6; TRAF6 FL; full-length TRAF6; Y2H; yeast two-hybrid system; RU; resonance unit; influenza virus; innate immune response

Domaines: Immunologie; Bactériologie; Virologie

Olivier Leymarie, Léa Meyer, Lionel Tafforeau, Vincent Lotteau, Bruno da Costa, et al.. Influenza virus protein PB1-F2 interacts with CALCOCO2 (NDP52) to modulate innate immune response. Journal of General Virology, 2017, 98 (6), pp.1196-1208. ⟨10.1099/jgv.0.000782⟩. ⟨hal-01911416⟩